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Background: Fibroblast growth factor 19 (FGF19) is produced in the small intestine and is involved in suppression of hepatic bile acid (BA) synthesis. FGF19 is also expressed in the liver and serum levels are elevated in adults with cholestatic liver disease. This may reflect a rescue mechanism to dampen liver injury caused by increased intrahepatic BAs., Objectives: To examine circulating FGF19 at early stages of biliary atresia and at short-term follow-up post-Kasai portoenterostomy (KPE) in relation to noncholestatic infants. The relationship between FGF19, BAs and markers for BA synthesis and hepatic gene expression of factors involved in BA metabolism were also evaluated., Methods: Liver tissue, portal and peripheral blood samples were obtained from fifteen patients at KPE; additional blood was collected 4-6 months after surgery. Two control groups were included; to examine possible changes related to surgery and to compare FGF19 in biliary atresia to noncholestatic infants., Results: Circulating FGF19 levels correlated to its hepatic gene expression at time of KPE in biliary atresia and levels were elevated compared to noncholestatic infants. At follow-up, FGF19 levels were markedly reduced, and the decline coincided with reductions in bilirubin and conjugated chenodeoxycholic acid and with increased levels of the BA synthesis marker C4., Conclusion: Elevated circulating FGF19 in biliary atresia is of hepatic origin and reduced following KPE. Changes in serum FGF19 may reflect the level of restoration of the enterohepatic circulation, and this warrants further long-term studies on the role of FGF19 in the cholestatic liver., (© 2020 The Association for the Publication of the Journal of Internal Medicine.)

Compared with humans, rodents have higher synthesis of cholesterol and bile acids (BAs) and faster clearance and lower levels of serum LDL-cholesterol. Paradoxically, they increase BA synthesis in response to bile duct ligation (BDL). Another difference is the production of hydrophilic 6-hydroxylated muricholic acids (MCAs), which may antagonize the activation of FXRs, in rodents versus humans. We hypothesized that the presence of MCAs is key for many of these metabolic differences between mice and humans. We thus studied the effects of genetic deletion of the Cyp2c70 gene, previously proposed to control MCA formation. Compared with WT animals, KO mice created using the CRISPR/Cas9 system completely lacked MCAs, and displayed >50% reductions in BA and cholesterol synthesis and hepatic LDL receptors, leading to a marked increase in serum LDL-cholesterol. The doubling of BA synthesis following BDL in WT animals was abolished in KO mice, despite extinguished intestinal fibroblast growth factor ( Fgf )15 expression in both groups. Accumulation of cholesterol-enriched particles ("Lp-X") in serum was almost eliminated in KO mice. Livers of KO mice were increased 18% in weight, and serum markers of liver function indicated liver damage. The human-like phenotype of BA metabolism in KO mice could not be fully explained by the activation of FXR-mediated changes. In conclusion, the presence of MCAs is critical for many of the known metabolic differences between mice and humans. The Cyp2c70 -KO mouse should be useful in studies exploring potential therapeutic targets for human disease., (Copyright © 2020 Straniero et al.)

Background & Aims: Disturbances of the enterohepatic circulation of bile acids (BAs) are seen in a number of clinically important conditions, including metabolic disorders, hepatic impairment, diarrhea, and gallstone disease. To facilitate the exploration of underlying pathogenic mechanisms, we developed a mathematical model built on quantitative physiological observations across different organs., Methods: The model consists of a set of kinetic equations describing the syntheses of cholic, chenodeoxycholic, and deoxycholic acids, as well as time-related changes of their respective free and conjugated forms in the systemic circulation, the hepatoportal region, and the gastrointestinal tract. The core structure of the model was adapted from previous modeling research and updated based on recent mechanistic insights, including farnesoid X receptor-mediated autoregulation of BA synthesis and selective transport mechanisms. The model was calibrated against existing data on BA distribution and feedback regulation., Results: According to model-based predictions, changes in intestinal motility, BA absorption, and biotransformation rates affected BA composition and distribution differently, as follows: (1) inhibition of transintestinal BA flux (eg, in patients with BA malabsorption) or acceleration of intestinal motility, followed by farnesoid X receptor down-regulation, was associated with colonic BA accumulation; (2) in contrast, modulation of the colonic absorption process was predicted to not affect the BA pool significantly; and (3) activation of ileal deconjugation (eg, in patents with small intestinal bacterial overgrowth) was associated with an increase in the BA pool, owing to higher ileal permeability of unconjugated BA species., Conclusions: This model will be useful in further studying how BA enterohepatic circulation modulation may be exploited for therapeutic benefits., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Background and Objectives: Bile acids (BAs) traversing the enterohepatic circulation (EHC) influence important metabolic pathways. By determining individual serum BAs in relation to markers of metabolic activity, we explored how diurnal variations in their EHC relate to hepatic metabolism in normal humans., Methods: Serum BAs, fibroblast growth factor 19 (FGF19), lipoproteins, glucose/insulin and markers of cholesterol and BA syntheses were monitored for 32 h in 8 healthy males. Studies were conducted at basal state and during initiation of cholestyramine treatment, with and without atorvastatin pretreatment. Time series cross-correlation analysis, Bayesian structural model and Granger causality test were applied., Results: Bile acids synthesis dominated daytime, and cholesterol production at night. Conjugated BAs peaked after food intake, with subsequent FGF19 elevations. BA synthesis was reduced following conjugated BA and FGF19 peaks. Cholestyramine reduced conjugated BAs and FGF19, and increased BA and cholesterol production; the latter effects attenuated by atorvastatin. The relative importance of FGF19 vs. conjugated BAs in this feedback inhibition could not be discriminated. Unconjugated BAs displayed one major peak late at night/early morning that was unrelated to FGF19 and BA synthesis, and abolished by cholestyramine. The normal suppression of serum triglycerides, glucose and insulin observed at night was attenuated by cholestyramine., Conclusions: Conjugated and unconjugated BAs have asynchronous rhythms of EHC in humans. Postprandial transintestinal flux of conjugated BAs increases circulating FGF19 levels and suppresses BA synthesis. Unconjugated BAs peak late at night, indicating a non-postprandial diurnal change in human gut microflora, the physiological implications of which warrants further study., (© 2018 The Authors. Journal of Internal Medicine published by John Wiley & Sons Ltd on behalf of Association for Publication of The Journal of Internal Medicine.)

Bile acid (BA) synthesis is regulated through suppression of hepatic cholesterol 7α-hydroxylase via farnesoid X receptor (FXR) activation in hepatocytes and/or enterocytes; in enterocytes, this process requires FGF19 signaling. To study these pathways, we quantified markers of BA synthesis (7α-hydroxy-4-cholesten-3-one [C4]) and cholesterol production (lathosterol), fibroblast growth factor (FGF)19, and BAs in serum from healthy male volunteers given 1 oral dose of the nonsteroidal FXR agonist Px-102 (0.15 mg/kg, 0.3 mg/kg, 0.6 mg/kg, 1.12 mg/kg, 2.25 mg/kg, 3.38 mg/kg, or 4.5 mg/kg). After 8 hours, serum levels of C4 decreased by 80% in volunteers given 0.15 mg/kg, whereas serum levels of FGF19 were unchanged. Serum levels of FGF19 increased significantly, in a dose-dependent manner, in volunteers given >0.3 mg/kg Px-102, up to as much as 1600%, whereas C4 levels remained significantly reduced (by >80%). For all doses, FGF19 levels returned to normal 24 hours after administration of Px-102. Serum levels of C4 decreased before levels of FGF19 levels increased, and were still reduced by 95% 24 hours after the highest dose (4.5 mg/kg) of Px-102, even though levels of FGF19 had returned to baseline. Our findings indicate that activation of hepatic FXR is able to suppress BA synthesis, independent of FGF19., (Copyright © 2018 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Previous studies have indicated that dietary intake of sugar may lower bile acid production, and may promote cholesterol gallstone formation in humans. We studied the influence of dietary sucrose on cholesterol and bile acid metabolism in the rat. In two different experiments, rats received high-sucrose diets. In the first, 60% of the weight of standard rat chow was replaced with sucrose (high-sucrose diet). In the second, rats received a diet either containing 65% sucrose (controlled high-sucrose diet) or 65% complex carbohydrates, in order to keep other dietary components constant. Bile acid synthesis, evaluated by measurements of the serum marker 7-alpha-hydroxy-4-cholesten-3-one (C4) and of the hepatic mRNA expression of Cyp7a1, was markedly reduced by the high-sucrose diet, but not by the controlled high-sucrose diet. Both diets strongly reduced the hepatic - but not the intestinal - mRNA levels of Abcg5 and Abcg8. The differential patterns of regulation of bile acid synthesis induced by the two sucrose-enriched diets indicate that it is not sugar per se in the high-sucrose diet that reduces bile acid synthesis, but rather the reduced content of fiber or fat. In contrast, the marked reduction of hepatic Abcg5/8 observed is an effect of the high sugar content of the diets., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Reduced plasma LDL-cholesterol is a hallmark of hyperthyroidism and is caused by transcriptional stimulation of LDL receptors in the liver. Here, we investigated whether thyroid hormone (TH) actions involve other mechanisms that may also account for the reduction in LDL-cholesterol, including effects on proprotein convertase subtilisin/kexin type 9 (PCSK9) and bile acid synthesis. Twenty hyperthyroid patients were studied before and after clinical normalization, and the responses to hyperthyroidism were compared with those in 14 healthy individuals after 14 days of treatment with the liver-selective TH analog eprotirome. Both hyperthyroidism and eprotirome treatment reduced circulating PCSK9, lipoprotein cholesterol, apoB and AI, and lipoprotein(a), while cholesterol synthesis was stable. Hyperthyroidism, but not eprotirome treatment, markedly increased bile acid synthesis and reduced fibroblast growth factor (FGF) 19 and dietary cholesterol absorption. Eprotirome treatment, but not hyperthyroidism, reduced plasma triglycerides. Neither hyperthyroidism nor eprotirome treatment altered insulin, glucose, or FGF21 levels. TH reduces circulating PSCK9, thereby likely contributing to lower plasma LDL-cholesterol in hyperthyroidism. TH also stimulates bile acid synthesis, although this response is not critical for its LDL-lowering effect., (Copyright © 2014 by the American Society for Biochemistry and Molecular Biology, Inc.)

Bile acids are synthesized from cholesterol in the liver and further metabolized by the gut microbiota into secondary bile acids. Bile acid synthesis is under negative feedback control through activation of the nuclear receptor farnesoid X receptor (FXR) in the ileum and liver. Here we profiled the bile acid composition throughout the enterohepatic system in germ-free (GF) and conventionally raised (CONV-R) mice. We confirmed a dramatic reduction in muricholic acid, but not cholic acid, levels in CONV-R mice. Rederivation of Fxr-deficient mice as GF demonstrated that the gut microbiota regulated expression of fibroblast growth factor 15 in the ileum and cholesterol 7α-hydroxylase (CYP7A1) in the liver by FXR-dependent mechanisms. Importantly, we identified tauro-conjugated beta- and alpha-muricholic acids as FXR antagonists. These studies suggest that the gut microbiota not only regulates secondary bile acid metabolism but also inhibits bile acid synthesis in the liver by alleviating FXR inhibition in the ileum., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Objective: Cholesterol and lipoprotein metabolism display pronounced gender differences. Premenopausal women have lower LDL and higher HDL cholesterol, whereas men display higher synthetic rates of bile acids and cholesterol. The effects of the administration of exogenous hormones to humans and animals indicate that these gender differences can often be explained by estrogens. We evaluated how increased levels of endogenous estrogens modulate cholesterol and lipoprotein metabolism in women., Methods and Results: We studied healthy women during initiation of in vitro fertilization using blood samples obtained when endogenous estrogens were low and high. Cholesterol in VLDL and LDL, but not in HDL, was reduced 20% when estrogens were high. Apolipoprotein B levels decreased 13%. Apolipoprotein A-I and triglyceride levels increased 8% and 37%, respectively, whereas lipoprotein(a) levels were unchanged. Circulating PCSK9, a suppressor of LDL receptors, was reduced 14% when estrogens were high. Serum markers of bile acid and cholesterol synthesis were unaltered. Growth hormone levels increased 3-fold when estrogens were high, whereas insulin-like growth factor-1 and fibroblast growth factor-21 concentrations were unaltered., Conclusion: In women, Apolipoprotein B-containing particles and circulating PCSK9 are reduced when endogenous estrogens are high, indicating that endogenous estrogens induce hepatic LDL receptors partly through a posttranscriptional mechanism. However, estrogens do not stimulate bile acid or cholesterol synthesis.

Background: Bile acid (BA) synthesis is essential in cholesterol and lipid homoeostasis., Methods: Serum samples from 435 normal and 23 cholecystectomized subjects were obtained after overnight fasting and assayed for markers of BA and cholesterol synthesis, as well as cholesterol absorption. We determined whether BA synthesis was related to fibroblast growth factor 19 (FGF19; a circulating metabolic regulator that is thought to inhibit BA synthesis), gender, age and serum lipids., Results: Bile acid synthesis varied more than 9-fold in normal individuals and was 29% higher in men than in women. Whilst low-density lipoprotein cholesterol increased with age, BA and cholesterol synthesis were stable. BA production was positively correlated with serum triglycerides (TGs), and 35% of individuals with a high level (>95th percentile) of BA synthesis had hypertriglyceridaemia (HTG) (>95th percentile). Serum FGF19 levels varied by 7-fold in normal individuals and were related inversely to BA synthesis but were not related to gender, plasma lipids or history of cholecystectomy., Conclusions: Bile acid synthesis has a wide inter-individual variation, is lower in women than in men and is correlated positively with serum TGs. High BA production is frequently linked to HTG. Age-related hypercholesterolaemia is not associated with changes in BA or cholesterol production, nor to an increase in cholesterol absorption. In humans, the circulating level of FGF19 may regulate hepatic BA production under fasting conditions., (2011 The Association for the Publication of the Journal of Internal Medicine.)

Atherosclerotic cardiovascular disease is a major problem despite the availability of drugs that influence major risk factors. New treatments are needed, and there is growing interest in therapies that may have multiple actions. Thyroid hormone modulates several cardiovascular risk factors and delays atherosclerosis progression in humans. However, use of thyroid hormone is limited by side effects, especially in the heart. To overcome this limitation, pharmacologically selective thyromimetics that mimic metabolic effects of thyroid hormone and bypass side effects are under development. In animal models, such thyromimetics have been shown to stimulate cholesterol elimination through LDL and HDL pathways and decrease body weight without eliciting side effects. We report here studies on a selective thyromimetic [KB2115; (3-[[3,5-dibromo-4-[4-hydroxy-3-(1-methylethyl)-phenoxy]-phenyl]-amino]-3-oxopropanoic acid)] in humans. In moderately overweight and hypercholesterolemic subjects KB2115 was found to be safe and well tolerated and elicited up to a 40% lowering of total and LDL cholesterol after 14 days of treatment. Bile acid synthesis was stimulated without evidence of increased cholesterol production, indicating that KB2115 induced net cholesterol excretion. KB2115 did not provoke detectable effects on the heart, suggesting that the pharmacological selectivity observed in animal models translates to humans. Thus, selective thyromimetics deserve further study as agents to treat dyslipidemia and other risk factors for atherosclerosis.

Plasma cholesterol increases in normal aging in both rodents and humans. This is associated with reduced elimination of cholesterol as bile acids (BAs) and decreased receptor-mediated clearance of plasma LDL, changes that can be reversed by treatment with growth hormone (GH). The level of intestinal absorption of cholesterol may also contribute to the development of hypercholesterolemia. In this study, we investigated whether cholesterol absorption increases with age and whether any such age-related change could be influenced by treatment with GH or ezetimibe (EZE). Male rats aged 6 and 18 mo were studied with and without GH or EZE treatment. BA synthesis was reduced and plasma cholesterol was increased in the old animals, whereas cholesterol absorption was unaltered. Cholesterol absorption was not altered by GH treatment but was reduced by EZE in both groups of animals. Hepatic LDL receptors (LDLRs), scavenger receptor class B type 1, and proprotein convertase subtilisin/kexin type 9 serine protease (PCSK9) transcripts were unchanged in old animals. GH treatment induced LDLRs, PCSK9 transcripts, and BA synthesis. We conclude that the age-induced hypercholesterolemia in the rat and its reversal by GH treatment relates to altered degradation of cholesterol in the liver and is not due to changes in cholesterol absorption.

Modulation of bile acid synthesis in human by cholestyramine or by chenodeoxycholic acid (CDCA) treatment affects lipoprotein metabolism leading to altered plasma lipid levels. The molecular changes caused by these treatments, which in turn influence lipoprotein metabolism, are still not entirely known in humans. In this study, mRNA levels were analyzed using real time RT-PCR in liver tissue from patients undergoing cholecystectomy due to gallstone disease. The patients were treated with either CDCA (n=6) or cholestyramine (n=5) for three weeks prior to surgery, six patients received no treatment and served as controls. Cholestyramine increased the expression of the LDL receptor (LDLR) by about 65% and that of proprotein convertase subtilisin kexin 9 (PCSK9) by 70%. After CDCA the levels of both LDLR and hydroxy-methyl-glutaryl coenzyme A reductase mRNA decreased approximately by 50%. The expression of PCSK9 was not changed. The mRNA levels of PCSK9, LDLR, and HMGCoAR were significantly correlated to those of sterol regulatory element binding protein 2 (SREBP2), indicating that SREBP2 is of importance in the regulation of the expression of these genes also in human liver.

Bile acids (BAs) traversing the enterohepatic circulation exert several important metabolic effects. Their hepatic synthesis, controlled by the enzyme cholesterol 7alpha-hydroxylase (CYP7A1), has a unique diurnal variation in man. Here we provide evidence that the transintestinal flux of BAs regulates serum levels of intestinal fibroblast growth factor 19 (FGF19) that in turn modulate BA production in human liver. Basal FGF19 levels varied by 10-fold in normal subjects, and were reduced following treatment with a BA-binding resin and increased upon feeding the BA chenodeoxycholic acid. Serum FGF19 levels exhibited a pronounced diurnal rhythm with peaks occurring 90-120 min after the postprandial rise in serum BAs. The FGF19 peaks in turn preceded the declining phase of BA synthesis. The diurnal rhythm of serum FGF19 was abolished upon fasting. We conclude that, in humans, circulating FGF19 has a diurnal rhythm controlled by the transintestinal BA flux, and that FGF19 modulates hepatic BA synthesis. Through its systemic effects, circulating FGF19 may also mediate other known BA-dependent effects on lipid and carbohydrate metabolism.

Background & Aims: The conversion of cholesterol to bile acids by the liver is an important regulator of body cholesterol homeostasis. In rodents, both cholesterol and bile acid synthesis have marked diurnal rhythms that peak synchronously at midnight. The aim of this study was to establish whether such diurnal rhythms are also present in healthy humans., Methods: Serum levels of the markers 7alpha-hydroxy-4-cholesten-3-one (C4) monitoring bile acid biosynthesis and lathosterol reflecting cholesterol synthesis were determined at 90-minute intervals in 8 human volunteers during standardized dietary conditions., Results: Serum C4 showed 2 distinct peaks (2- to 4-fold above baseline) during a 24-hour period, the first at 1:00 pm and the second at 9:00 pm. During the night, C4 levels declined, and they returned to baseline levels the next morning. In contrast, serum lathosterol levels peaked at night, between midnight and 4:00 am. The diurnal changes of C4 were not synchronous with serum lipid changes or with the postprandial increase in serum bile acids and were maintained in cholecystectomized subjects., Conclusions: Bile acid synthesis in humans has a diurnal rhythm, with 2 peaks during the daytime, that is opposite from the circadian rhythm of cholesterol synthesis. This is completely different from the pattern in rodents and indicates the presence of an important species variation in the regulation of cholesterol homeostasis.

The maintenance of adequate amounts of bile acids in the liver, biliary tract, and intestine requires a finely tuned control of their synthesis. A paper in this issue of Cell Metabolism by Inagaki et al. (2005) indicates that sensing of the levels of bile acids in the intestine may trigger the secretion of a hormone which regulates bile acid production in the liver.

Transcriptional regulation of the cholesterol 7alpha-hydroxylase (CYP7AI) gene is of critical importance for bile acid and cholesterol metabolism. We evaluated the physiological significance of two common polymorphisms (-203C/A and -469T/C) in the promoter region of the CYP7AI gene. No evidence was found for physiological differences between either the -203C and -203A alleles or the -469T and -469C alleles in transient transfection studies using native 834bp promoter constructs. Moreover, no association was observed between the CYP7AI promoter polymorphisms and the hepatic cholesterol 7alpha-hydroxylase activity and parameters of bile acid synthesis rates, as analyzed in subjects with gallstone disease. In addition, no relationships were found between the promoter polymorphisms and plasma LDL cholesterol concentration in association studies conducted in three different groups of middle-aged Swedish men. Finally, near complete allelic association was found between the two promoter polymorphisms and the IVS6+363G/A polymorphism at the 3' end of the CYP7AI gene (|D'|=0.98), indicating strong linkage disequilibrium across the whole CYP7AI gene. It is concluded that common polymorphisms of the CYP7A1 gene do not contribute to variation in cholesterol 7alpha-hydroxylase activity, rates of bile acid synthesis and plasma LDL cholesterol concentration in humans.

Previous studies have established that growth hormone (GH) has many important effects on the regulation of cholesterol and lipoprotein metabolism. However, human GH (hGH) can also bind to prolactin receptors, eliciting prolactin receptor-mediated effects. In this study, we evaluated whether hGH can exert such responses in currently used animal models and whether prolactin affects lipoprotein and/or hepatic cholesterol metabolism. Normal and hypophysectomized (Hx) male rats were given either hGH or bovine GH, the latter unable to bind to the prolactin receptor. The hormones were continuously infused by use of subcutaneous osmotic mini-pumps for 7 days; blood and livers were collected after euthanasia. Both hormones stimulated hepatic LDL receptor expression and bile acid synthesis to a similar extent and normalized the altered plasma lipoprotein pattern in Hx rats. Prolactin, injected twice daily to Hx male rats, did not exert any effects on the plasma lipoprotein pattern or on cholesterol metabolism. We conclude that previously established effects of hGH on cholesterol metabolism are not mediated by prolactin in male rats.

Objective: Growth hormone (GH) induces hepatic low-density lipoprotein (LDL) receptors and lowers plasma cholesterol. We characterized the influence of GH treatment on plasma LDL clearance in normal humans and investigated the relative role of LDL receptor (LDLR) activity and stimulation of bile acid synthesis in subjects with different LDLR expression., Methods and Results: Plasma clearance of autologous 125I-LDL was measured before and during 3 weeks of treatment with GH (0.1 IU/kg per day) in 9 healthy young males. Plasma LDL cholesterol was reduced by 13% and the fractional catabolic rate of LDL increased by 27%. More marked changes were seen in a patient with hypopituitarism substituted with GH (0.07 IU/kg per day) for 3 months. In a second study, GH dose-dependently reduced LDL cholesterol and increased Lp(a) levels in 3 groups of males: younger and elderly healthy subjects and heterozygous familial hypercholesterolemia (FH). No effect on bile acid synthesis measured by the plasma marker 7alpha-hydroxy-4-cholesten-3-one was observed. In an LDLR-deficient FH homozygote, LDL cholesterol was not affected by GH., Conclusions: GH treatment reduces plasma LDL cholesterol by inducing LDL clearance. In humans, LDLR expression is a prerequisite for this effect, whereas it is not related to stimulation of bile acid synthesis.

We describe an accurate method for monitoring the enzymatic activity of hepatic cholesterol 7alpha-hydroxylase (C7alphaOH; CYP7A1), the rate-limiting and major regulatory enzyme in the synthesis of bile acids. Assay of 7alpha-hydroxy-4-cholesten-3-one (C4), an intermediate in bile acid synthesis, revealed that the level of C4 in peripheral blood serum or plasma showed a strong correlation to the enzymatic activity of hepatic C7alphaOH, both at steady-state conditions (r = 0.929) as well as during the rapid changes that occur during the diurnal phases. This assay should be of value in clarifying the regulation of bile acid synthesis in vivo in laboratory animals and humans since it allows for the monitoring of hepatic C7alphaOH activity using peripheral blood samples.

It has been reported that there is a coordinate regulation of sterol 27-hydroxylase (CYP27A1) and cholesterol 7alpha-hydroxylase (CYP7A1) in rats. Thus, the levels of the mRNA corresponding to these two enzymes were found to change in the same direction in rat liver and in isolated rat hepatocytes. In contrast, other groups have not seen such regulation of CYP27A1 in rabbit liver or in rat liver when using an activity assay. In the present work, the effect of bile acid treatment on human CYP27A1/luciferase reporter activity was studied in a transient transfection assay in human liver-derived HepG2 cells. Neither the endogenous 27-hydroxylase activity nor the CYP27A1/luciferase reporter activity were down-regulated by treatment of HepG2 cells with chenodeoxycholic acid or taurochenodeoxycholic acid. We also measured CYP27A1 mRNA and CYP7A1 mRNA in liver of humans subjected to treatment with chenodeoxycholic acid, ursodeoxycholic acid, hydroxymethylglutaryl (HMG)-CoA reductase inhibitor and a combination of HMG-CoA reductase inhibitor and cholestyramine. There was a 60-fold variation in the levels of CYP7A1 mRNA but only a 5-fold variation in the levels of CYP27A1 mRNA. There was no correlation between the two mRNA species. It is concluded that, in humans, there is little or no coordinate regulation of CYP7A1 and CYP27A1 at the transcriptional level, and that CYP27A1 is not subject to a negative feedback control by bile acids. The results underline that marked species differences may exist in mechanisms for control of synthesis of bile acids and cholesterol homeostasis.

Familial hypertriglyceridemia (FHTG), a disease characterized by elevated plasma very low density lipoprotein triglyceride levels, has been associated with impaired intestinal absorption of bile acids. The aim of this study was to test the hypothesis that defects in the active ileal absorption of bile acids are a primary cause of FHTG. Single-stranded conformation polymorphism analysis was used to screen the ileal Na(+)/bile acid cotransporter gene (SLC10A2) for FHTG-associated mutations. Analysis of 20 hypertriglyceridemic patients with abnormal bile acid metabolism revealed 3 missense mutations (V98I, V159I, and A171S), a frame-shift mutation (646insG) at codon 216, and 4 polymorphisms in the 5' flanking sequence of SLC10A2. The SLC10A2 missense mutations and 5' flanking sequence polymorphisms were not correlated with bile acid production or turnover in the hypertriglyceridemic patients and were equally prevalent in the unaffected control subjects. In transfected COS cells, the V98I, V159I, and A171S isoforms all transported bile acids similar to the wild-type SLC10A2. The 646insG frame-shift mutation abolished bile acid transport activity in transfected COS cells but was found in only a single FHTG patient. These findings indicate that the decreased intestinal bile acid absorption in FHTG patients is not commonly associated with inherited defects in SLC10A2.

Background: Apolipoprotein (apo) A-I is the major protein component of HDL, a cholesterol transport particle that protects against atherosclerosis. Apo A-I is believed to promote reverse cholesterol transport, transferring cholesterol from peripheral cells to the liver for subsequent elimination. To test this hypothesis in humans, we measured fecal steroid excretion before and after the intravenous infusion of human proapo A-I (precursor of apo A-I) liposome complexes., Methods and Results: Four subjects with heterozygous familial hypercholesterolemia w re studied under standardized conditions. The fecal excretion of bile acids and neutral sterols was determined for 9 days before and 9 days after an intravenous infusion of recombinant human proapo A-I (4 g protein) liposome complexes. Plasma apoA-I and HDL cholesterol levels increased transiently (mean peak concentrations were 64% and 35% above baseline, respectively) during the first 24 hours. Mean lipoprotein lipid and apolipoprotein levels were not different during the 2 collecting periods, however. Serum lathosterol, a precursor of cholesterol whose concentration reflects the rate of cholesterol synthesis in vivo, was also unchanged. The fecal excretion of cholesterol (neutral sterols and bile acids) increased in all subjects (mean increase, +39% and +30%, respectively), corresponding to the removal of approximately 500 mg/d excess cholesterol after infusion. Control infusions with only liposomes in 2 of the patients did not influence lipoprotein pattern or cholesterol excretion., Conclusions: Infusion of proapoA-I liposomes in humans promotes net cholesterol excretion from the body, implying a stimulation of reverse cholesterol transport. This mechanism may prove useful in the treatment of atherosclerosis.

Based on an improved molecular understanding of how bile acid metabolism is regulated, an exciting period of research developments can be expected. By new ways of stimulating cholesterol breakdown to bile acids, novel therapeutic principles can be forseen which will further improve our potential for treating and preventing atherosclerosis.

Growth hormone (GH) has an important role in the regulation of hepatic LDL receptor expression and plasma lipoprotein levels. This investigation was undertaken to characterize the effects of GH on hepatic cholesterol and bile acid metabolism in the rat. In hypophysectomized (Hx) rats, the activities of the rate-limiting enzymes in cholesterol and bile acid biosynthesis, 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG CoA reductase) and cholesterol 7alpha-hydroxylase (C7alphaOH), were reduced by 71 and 64%, respectively. HMG CoA reductase mRNA levels were reduced by 37%, whereas C7alphaOH mRNA was increased by 81%. LDL receptor expression was reduced by 18% in Hx rats, without any change in the LDL receptor mRNA levels. Although the normal diurnal variation of C7alphaOH activity was preserved in Hx rats, the activity of C7alphaOH was much reduced both at midday and midnight. Total hepatic cholesterol was increased by 14% in Hx animals whereas microsomal cholesterol was unchanged. The rate of cholesterol esterification was enhanced (by 38%) in liver microsomes from Hx rats. Stepwise hormonal substitution of Hx rats showed that GH, but not thyroid hormone or cortisone, was essential to normalize the enzymatic activity of C7alphaOH. GH also normalized the altered plasma lipoprotein pattern in Hx rats, and increased the fecal output of bile acids. The latter effect was particularly evident when GH was combined with cortisone and thyroid hormone. Also in normal rats, GH stimulated C7alphaOH activity. In conclusion, GH has an essential role to maintain a normal enzymatic activity of C7alphaOH, and this, at least in part, explains the effects of GH on hepatic cholesterol metabolism. GH is also of critical importance to normalize the altered plasma lipoprotein pattern in Hx rats.

Aims/methods: Bile acid kinetics and biliary lipid composition were studied in seven patients, aged 17-70 years with cystic fibrosis. All patients were of normal height and weight, and were in good clinical condition. Ultrasonography indicated a small gallbladder in one and non-visualized gallbladder in two patients. Serum concentrations of cholesterol and transaminases were essentially normal. Substitution with pancreatic enzymes was discontinued at least 1 week before the investigation. Bile acid kinetics were determined by the isotope dilution technique using [24-14C] cholic and [24-14C] chenodeoxycholic acids., Results: The mean pool size of cholic acid was 3.3 (range 0.8-6.9) mmol, and that of chenodeoxycholic acid 2.3 (1.2-2.7) mmol, corresponding to 49 +/- 16 and 36 +/- 4 mumol/kg, respectively. The mean synthesis of cholic acid was 1.3 (0.5-3.6) mmol/day and of chenodeoxycholic acid 0.8 (0.2-1.7) mmol/ day and related to body weight 20 +/- 6 and 12 +/- 3 mumol.kg.day-1, respectively. Fractional turnover rates averaged 0.48 (0.24-0.67) and 0.36 (0.10-0.65) day-1, respectively. The kinetic values were not significantly different from controls, aged 21 to 68 years. The biliary lipid composition of fasting gallbladder bile showed a low-normal molar percentage of cholesterol, and in no case was bile supersaturated. The duodenal bile acid concentration was similar in patients and controls, but the bile acid distribution was significantly different; cholic acid constituted a higher percentage (p < 0.001) and chenodeoxycholic and deoxycholic acid lower percentages (p < 0.05 and p < 0.01, respectively) in cystic fibrosis patients than in controls., Conclusions: The findings of normal concentrations of bile acids in duodenal bile and normal to large pool sizes of bile acids in all patients, despite a marked fat malabsorption, are in contrast to some previous reports. The data indicate that biliary lipid metabolism is normal in well-nourished and well-controlled adult patients with cystic fibrosis.

A new model has been developed to characterise the effect of a standardised ileal exclusion on bile acid, cholesterol, and lipoprotein metabolism in humans. Twelve patients treated by colectomy and ileostomy for ulcerative colitis were studied on two occasions: firstly with a conventional ileostomy and then three months afterwards with an ileal pouch operation with an ileoanal anastomosis and a protective loop ileostomy, excluding on average 95 cm of the distal ileum. The ileostomy contents were collected during 96 hours and the excretion of bile acids and cholesterol was determined using gas chromatography-mass spectrometry. Fasting blood and duodenal bile samples were collected on two consecutive days. After the exclusion of the distal ileum, both cholic and chenodeoxycholic acid excretion in the ileostomy effluent increased four to five times without any change in cholesterol excretion. Serum concentrations of lathosterol (a marker of cholesterol biosynthesis) and 7 alpha-hydroxycholesterol (a marker for bile acid biosynthesis) were increased several fold. Plasma concentrations of total VLDL triglycerides were also increased whereas the concentrations of total and LDL cholesterol, and apolipoprotein B were decreased. There were no changes in biliary lipid composition or cholesterol saturation of bile. The results show that the exclusion of about 95 cm of distal ileum causes malabsorption of bile acids but apparently not of cholesterol. The bile acid malabsorption leads to increased synthesis of both bile acids and cholesterol in the liver. It is suggested that bile acids can regulate cholesterol synthesis by a mechanism independent of the effect of bile acids on cholesterol absorption. The enhanced demand for cholesterol also leads to a decrease in plasma LDL cholesterol and apolipoprotein B concentrations. The malabsorption of bile acids did not affect biliary lipid composition or cholesterol saturations of VLDL triglycerides.

The synthesis of bile acids is regulated by a homeostatic mechanism in which bile acids returning to the liver from the intestine inhibit their own synthesis. Serum concentrations of the bile acid intermediate 7 alpha-hydroxy-4-cholesten-3-one reflect the rate of bile acid synthesis whereas bile acid malabsorption can be determined by the SeHCAT test. This study was done to evaluate the correlation between the two tests in humans. Twenty eight patients with chronic diarrhoea were included in the study. Fasting serum was collected for the determination of 7 alpha-hydroxy-4-cholesten-3-one, and on the same day the gamma emitting bile acid analogue SeHCAT was given orally and its fractional catabolic rate assessed by repeated external counting over the upper abdomen during the next seven days. There was a highly significant positive correlation between the two tests (Rs = 0.80, p < 0.001). The results show a close relation between intestinal loss and hepatic synthesis of bile acids and imply that analysis of 7 alpha-hydroxy-4-cholesten-3-one in serum should now be evaluated as a possible convenient method for assessing bile acid malabsorption in patients with diarrhoea.

The time required for precipitation of cholesterol crystals (nucleation time, NT) was determined and related to the cholesterol saturation in gall bladder bile of gall stone free subjects (n = 11), patients with pigment stones (n = 3), and patients with cholesterol gall stones (n = 30) undergoing cholecystectomy. Seven of the gall stone patients had been treated with chenodeoxycholic acid (CDCA) and nine with ursodeoxycholic acid (UDCA), 15 mg/kg/day for three weeks before operation. NT was longer in gall stone free subjects (mean, 20 days), patients with pigment stones (14 days) and patients treated with CDCA (24 days) and UDCA (17 days) compared with untreated patients with cholesterol gall stones (1.5 days). In spite of low cholesterol saturation and prolonged NT, and in contrast to those treated with CDCA, four of the nine patients treated with UDCA had cholesterol crystals in their bile. These observations give further support to the concept that the mechanism for inducing gall stone dissolution may be different for CDCA and UDCA.

The effects of urodeoxycholic acid on ileal absorption of bile acids and on serum bile acid and lipoprotein concentrations were studied. Eight healthy subjects were investigated. The gamma emitting bile acid analogue, SeHCAT, was given orally and its fractional catabolic rate and seven day retention were assessed by repeated external counting over the upper abdomen during the next seven days. Ursodeoxycholic acid was then given orally at a dose of 15 mg/kg/day for three weeks and the study was repeated during treatment. The fractional catabolic rate increased by 64% (mean (SD), 0.333 (0.159) v 0.203 (0.061)/day; p less than 0.05) and seven day retention decreased by 44% (15(10) v 27(10)%, p less than 0.001), indicating bile acid malabsorption. Total serum cholesterol fell from 5.79 (1.22) to 5.50 (1.18) mmol/l (p = 0.05), while serum ursodeoxycholic acid increased 22 fold (7.87 (2.67) v 0.34 (0.24) mumol/l, p less than 0.001). Five of the subjects continued taking 30 mg/kg/day of ursodeoxycholic acid for one week and showed an increase in fractional catabolic rate of 81% (0.300 (0.091) v 0.166 (0.037)/day; p less than 0.05) and a fall in seven day retention of 50% (16 (12) v 32 (8)%, p less than 0.01). There were significant reductions in total cholesterol (5.36 (1.71) v 6.08 (1.47) mmol/l; p less than 0.05) and low density lipoprotein cholesterol (3.70 (1.33) v 4.58 (1.16) mmol/l; p less than 0.05). The results support the concept tht ursodeoxycholic acid treatment interferes with the absorption of endogenous bile acids, and emphasise the beneficial effects of this treatment of lipoprotein concentrations in man.

The activity of acyl CoA: cholesterol acyltransferase (ACAT), which catalyzes the esterification of cholesterol, was studied in liver microsomes obtained from cholestyramine-treated gallstone patients (n = 12) and patients with Crohn's disease who had undergone partial ileal resection (n = 11). Gallstone patients (n = 33) and gallstone-free subjects undergoing cholecystectomy because of polyps of the gallbladder (n = 8) served as controls. The mean levels of the ACAT activity were the same in the gallstone and the gallstone-free patient groups (6.0 +/- 0.4 and 6.1 +/- 1.1 pmol/min per mg protein, respectively). When exogenous cholesterol was added to the assay system the activities were increased four- to fivefold in both groups. The ACAT activity tended to be increased in the cholestyramine-treated patients (8.1 +/- 1.8 pmol/min per mg protein), and was significantly enhanced (P less than 0.005) in the ileal-resected patients (12.3 +/- 2.3 pmol/min per mg protein). When the enzyme activity was determined with added exogenous cholesterol, it was significantly higher compared to the controls in both the cholestyramine-treated patients and the patients with ileal resection (57.9 +/- 11.6 and 50.0 +/- 10.3 pmol/min per mg protein, respectively). The content of free and esterified cholesterol in liver homogenates and microsomes was not significantly different between the patient groups. We conclude that ACAT activity is increased in patients with interruption of the enterohepatic circulation of bile acids, and speculate that this reflects a stimulated uptake of lipoprotein cholesterol and may indicate that more cholesteryl esters are incorporated into very low density lipoproteins.

Interruption of the enterohepatic circulation of bile acids by cholestyramine or colestipol influences the hepatic metabolism of cholesterol in many ways. The synthesis of bile acids is increased, as reflected by a several-fold increase in the activity of the cholesterol 7 alpha hydroxylase, the rate-determining enzyme in bile acid synthesis. The increased metabolism of cholesterol to bile acids causes an enhanced demand of cholesterol in the hepatocytes which respond with both new synthesis of cholesterol, as reflected in a several-fold increase of the HMG-CoA reductase activity, and increased expression of LDL receptors. As a consequence, the plasma level of LDL-cholesterol is lowered. The hepatic secretion rate of VLDL particles is increased. Cholestyramine therapy does not affect the output of biliary lipids or the cholesterol saturation of bile, indicating that treatment with bile acid sequestrants should not be associated with any increased risk of gallstone formation.

Bile acid kinetics and biliary lipid composition were determined in seven subjects with primary dysbetalipoproteinaemia. They were all homozygous for the apolipoprotein E isoform E-2 and six of them were hyperlipidaemic (type III hyperlipoproteinaemia). With or without hyperlipidaemia, the apo E-2/2 phenotype was associated with increased bile acid formation (mean increase compared with 32 normolipidaemic controls, 43%; P less than 0.025). The biliary lipid composition was not different from that seen in the controls. The results indicate that the uptake by the liver of apo E-containing remnant particles is of importance for the regulation of hepatic cholesterol metabolism in man. It is suggested that hepatic cholesterol synthesis is stimulated in dysbetalipoproteinaemia, and that this leads to a compensatory increase in bile acid synthesis.

Supersaturation of bile with cholesterol predisposes to the development of cholesterol gallstones. To identify the factors determining cholesterol saturation of bile, we analyzed the lipid composition of stimulated duodenal bile in 60 healthy subjects of various ages (31 men and 29 women) who were not obese and were free of gallstones. A positive correlation between age and cholesterol saturation of bile was found (P less than 0.001). To analyze the relation between age and cholesterol saturation, we studied the rates of hepatic secretion of biliary lipids and the kinetics of cholic acid and chenodeoxycholic acid in 22 and 18 of the subjects, respectively. Age was positively correlated with the cholesterol secretion rate (r = 0.48) and negatively correlated with bile acid synthesis (r = -0.60) and the size of the cholic acid pool (r = -0.54). We conclude that cholesterol saturation of bile increases with age as a consequence of enhanced hepatic secretion of cholesterol and decreased bile acid synthesis. These findings may explain why age is a risk factor for the development of cholesterol gallstones.

Cholesterol in plasma high density lipoproteins (HDL) has been proposed to serve as preferential precursor for bile acid biosynthesis in the liver. Furthermore, a negative relationship between plasma levels of HDL cholesterol and biliary saturation with cholesterol has been reported in healthy females. We have performed metabolic studies on a female patient with fish eye disease, a familial condition where plasma HDL levels are reduced by 90% and the concentration of plasma triglycerides is moderately increased. Both the total production of bile acids and the net steroid 'balance' (reflecting total body cholesterol synthesis) were within the range seen in normolipidaemic as well as in hypertriglyceridaemic females. Also the biliary lipid composition and cholesterol saturation of bile were normal. A qualitative abnormality in the bile acid pattern was observed, however, in that the ratio between the synthesis of cholic acid and chenodeoxycholic acid was reduced. It is concluded that a low HDL cholesterol level is not necessarily associated with quantitative abnormalities of biliary lipid metabolism. The abnormal bile acid synthesis ratio may reflect changes in the hepatic precursor pools of cholesterol as the consequence of HDL deficiency, however.

Bile acid and plasma endogenous triglyceride kinetics were determined under standardized dietary conditions in 47 hyperlipidemic subjects with the aid of [14C]cholic acid, [14C]chenodeoxycholic acid, and [3H]glycerol, respectively. On the basis of their lipoprotein pattern the patients were separated into three groups characterized by hyperlipoproteinemia (HLP) type IIa (n = 19), type IIb (n = 6), and type IV (n = 22). In keeping with previous reports from this laboratory the total bile acid formation reports from this laboratory the total bile acid formation in HLP type IV (19.5 +/- 2.2) mumol kg-1d-1, mean +/- SEM) exceeded that encountered in type IIa (10.7 +/- 0.9 mumol kg-1d-1, P less than 0.005). This difference was mainly due to an increased synthesis of cholic acid in type IV HLP (12.7 +/- 1.7 mumol kg-1d-1 vs. 6.1 +/- 0.5 mumol kg-1d-1, P less than 0.005). Bile acid formation in type IIb HLP was essentially within the limits recorded for type IIa. Apparent plasma triglyceride formation (as calculated from the 10-hr radioactivity decay curve) averaged 10.5 +/- 0.7 mumol kg-1hr-1 in type IIa HLP and was significantly higher in type IIb (20.7 +/- 1.9 mumol kg-1hr-1, P less than 0.001) and in type IV (22.1 +/- 1.4 mumol kg-1hr-1, P less than 0.001). The apparent fractional turnover rate of plasma triglyceride in type IV HLP (0.147 +/- 0.011 hr-1) was lower than that encountered in type IIa (0.188 +/- 0.008, P less than 0.01) and in type IIb (0.177 +/- 0.011 hr-1). The apparent production of plasma triglycerides and the formation of cholic acid correlated in type IIa (r = +0.69, P less than 0.001) and in type IV HLP (r = +0.70, P less than 0.001). A similar pattern was seen for total bile acid formation, while chenodeoxycholic acid showed a correlation to apparent triglyceride synthesis only in type IV HLP. It is suggested that an increased formation of plasma triglycerides--monitoring very low density lipoprotein synthesis--is linked to an enhanced degradation of cholesterol to bile acids and that there is an integrated regulation of the metabolism of these two parameters.

Combined gas-liquid chromatography-mass spectrometry with specific ion monitoring (mass fragmentography) has been used for assay of cholic acic (C), chenodeoxycholic acid (CD), and deoxycholic acid (D) in human serum. Deuterium-labeled C and D were used as internal standards. The relative standard deviation of duplicate samples was 3, 4, and 7% for C, CD, and D, respectively. The variation within the same individual in the fasting state was small, while the day-to-day variation was greater, especially for the dihydroxy bile acids. In normal control subjects (n = 24), the fasting serum concentration of C averaged 184 +/- 24 ng/ml (mean +/- SEM), and that of CD and D 526 +/- 62 and 407 +/- 44 ng/ml, respectively. Patients with type IIa hyperlipoproteinemia (n = 32) displayed low values of serum bile acids, with a C concentration of 121 +/- 11 ng/ml (P less than 0.01 vs. controls). A similar pattern was seen in patients with a type IIb lipoprotein pattern (n = 10). Subjects with type IV hyperlipoproteinemia (n = 32) showed serum bile acid levels within the normal limits. No relationship to age, sex, or body weight was seen in any of the patient subgroups. Bile acid kinetics were determined with an isotope dilution technique using 14C-labeled C and CD under steady state conditions in control subjects and patients with type IIa and type IV hyperlipoproteinemia. The serum concentration of C correlated significantly to its pool size in control subjects and in patients with type IIa hyperlipoproteinemia but not in patients with type IV. The serum level of CD was not related to CD pool size in any of the subgroups. The data obtained are discussed in relation to present concepts of the enterohepatic circulation. It is suggested that the intestinal content of C in the fasting state is proportional to the total C pool size. The possibility of a defective intestinal uptake of C in some patients with type IV hyperlipoproteinemia is raised.

1. The liver microsomal metabolism of [4-14C]cholesterol, endogenous cholesterol, 7 alpha-hydroxy-4-[6 beta-3H]cholesten-3-one, 5-beta-[7 beta-3H]cholestane-3 alpha, 7 alpha-diol and [3H]lithocholic acid was studdied in control and clofibrate (ethyl p-chlorophenoxyisobutyrate)-treated rats. 2. The extent of 7 alpha-hydroxylation of exogenous [414C]cholesterol and endogenous cholesterol, the latter determined with a mass fragmentographic technique, was the same in the two groups of rats. The extent of 12 alpha-hydroxylation of 7 alpha-hydroxy-4-cholesten-3-one and 5 beta-cholestane-3 alpha, 7 alpha-diol was increased by about 60 and 120% respectively by clofibrate treatment. The 26-hydroxylation of 5 beta-cholestane-3 alpha, 7 alpha-diol was not significantly affected by clofibrate. The 6 beta-hydroxylation of lithocholic acid was about 80% higher in the clofibrate-treated animals than in the controls. 3. The results are discussed in the context of present knowledge about the liver microsomal hydroxylating system and bile acid formation in patients with hypercholesterolaemia, treated with clofibrate.

The aim of this investigation was to study the influence of chenodeoxycholic acid administration on cholesterol and bile acid synthesis in germ-free rats. Seven rats were fed a basal diet and 2 groups of 4 rats received the same diet supplemented with 0.4 and 1% chenodeoxycholic acid, respectively. After 6 weeks, feces were collected in one 3- and one 4-day pool for analysis of cholesterol and bile acids. When the sampling period was finished, the rats were killed and the liver microsomal fractions isolated. The activities of HMG CoA reductase and cholesterol 7 alpha-hydroxylase were determined, the 7 alpha-hydroxylase by a mass fragmentographic method. The 2 dominating bile acids in the untreated rats were cholic acid and beta-muricholic acid. During treatment with chenodeoxycholic acid, 60--70% of this bile acid was converted into alpha- and beta-muricholic acid, indicating a high activity of the 6 beta-hydroxylase. The excretion of cholic acid was almost completely inhibited and the 7 alpha-hydroxylase activity was decreased ca 75% in the rats fed 1% chenodeoxycholic acid. The activity of the hepatic HMG CoA reductase as unchanged. The fecal excretion of cholesterol increased 2--3 times. An accumulation of cholesterol was seen in the rats treated with 1% chenodeoxycholic acid, which was probably a result of the decreased catabolism of cholesterol to bile acids.

The aim of the present study was to evaluate the diagnostic value of measuring individual serum bile acids in patients with suspected alcoholic liver disease. A highly accurate and specific mass-fragmentographic technique with high sensitivity was used. Anicteric patients with fatty liver (n = 10) and liver cirrhosis (n = 9) were compared with healthy controls (n = 27). The measurement of serum bile acids did not discriminate patients with fatty liver from controls. In general, an increased serum level of cholic acid indicated more serious liver disease. Determination of chenodeoxycholic acid and deoxycholic acid did not add any further diagnostic information.

The serum concentrations of cholic acid (C), chenodeoxycholic acid (CD), and deoxycholic acid (D) before and after a standardised meal were determined in five healthy female subjects using a highly specific and accurate gas chromatographic-mass spectrometric technique. The C level rose significantly 60 minutes after the meal, reached a peak after 90 minutes, and had returned to the original level after 150 minutes. In contrast, the serum concentrations of CD and D displayed a significant rise by 30 minutes, reached a peak after 90 minutes, but had not returned to fasting levels after 150 minutes. The serum bile acid responses after a meal suggest that there is considerable absorption of dihydroxy bile acids in the proximal small intestine in man.

Previous studies have indicated that quantitative as well as qualitative abnormalities of bile acid metabolism frequently occur in hypercholesterolaemia. In order to determine if this is a feature of familial hypercholesterolaemia, bile acid kinetics and biliary lipid composition were determined in 15 affected (heterozygous) and six unaffected siblings of four kindreds with familial hypercholesterolaemia. Furthermore, serum levels of cholic acid, chenodeoxycholic acid and deoxycholic acid were measured with a mass fragmentographic technique in 15 members of two of the kindreds, and secretion rates of biliary lipids were measured in six members of two kindreds. No differences with regard to these parameters between affected and unaffected siblings could be detected. There was a close resemblance between relatives of a given kindred concerning bile acid pool size and serum bile acid levels. No evidence for a defective bile acid metabolism in familial hypercholesterolaemia could be gained from the present study. It is concluded that the deficient receptor-mediated elimination of low density lipoprotein cholesterol in this disorder does not influence the maintenance of normal bile acid metabolism.

Kinetics of [24-14C] cholic acid and [24-14C] chenodeoxycholic acid and biliary lipid composition were determined in patients with chronic alcoholic pancreatitis with (n = 8) and without (n = 8) steatorrhoea. Pool sizes, syntheses and fractional catabolic rates of the two bile acids were not significantly different from corresponding values in healthy controls. Biliary lipid composition was normal in patients without steatorrhoea. Patients with steatorrhoea, on the other hand, had lower cholesterol saturation of bile than corresponding controls. This is probably because of malabsorption of cholesterol.

Two distinct lipoprotein receptors can be expressed in the dog liver. One is the apolipoprotein (apo-) B,E receptor. This receptor binds apo-B-containing low density lipoproteins (LDL), as well as apo-E-containing lipoproteins, such as the cholesterol-induced high density lipoproteins (HDL(c)). The second hepatic lipoprotein receptor is the apo-E receptor. It binds apo-E HDL(c) and chylomicron remnants, but not LDL. The present studies were undertaken to determine whether short-term (acute) regulation of the two receptors can occur in response to perturbations in hepatic cholesterol metabolism. The design used three groups of experimental animals: (a) immature dogs (with both hepatic apo-B,E and apo-E receptors expressed), (b) adult dogs (with predominantly the apo-E receptor expressed and little detectable apo-B,E receptor binding activity), and (c) dogs treated with the bile acid sequestrant cholestyramine or those that have undergone biliary diversion (with apo-E receptors and induced apo-B,E receptors). In the first series of experiments, changes in hepatic lipoprotein receptor expression were studied by delivering cholesterol to the liver via intestinal lymph lipoproteins. Dog lymph (5-11 mg of triglycerides/min per kg of body weight, 0.15-0.3 mg of cholesterol/min per kg) or saline were infused intravenously for 6-8 h into matched pairs of dogs. Serial liver biopsies were obtained at intervals of 1-2 h. A progressive loss of specific (calcium-dependent) binding of LDL was seen in hepatic membranes from both immature and cholestyramine-treated dogs. After 4-6 h of lymph infusion, almost no apo-B,E receptor binding could be detected. The decrease in binding of apo-E HDL(c) to the same membranes was much less pronounced, and could be explained by a loss of binding of HDL(c) to the apo-B,E receptor; there was little or no effect on apo-E receptor binding. In the second series of experiments, the effects of a diminished hepatic demand for cholesterol on lipoprotein receptor expression were studied by suppressing bile acid synthesis. The bile acid taurocholate (2-3 mumol/kg per min) was infused intravenously over a 6-h interval. This resulted in a progressive loss of LDL binding to liver membranes of immature or cholestyramine-treated dogs. The infusion of taurocholate for 6 h did not significantly alter the expression of the apo-E receptor binding activity, whereas apo-B,E receptor activity was rapidly down-regulated. Preparation of a bile fistula in adult dogs markedly induced the expression of the apo-B,E receptor. In this state, the binding activity of the apo-B,E receptor could be almost totally abolished by reinfusion of taurocholate for 6 h, without profoundly affecting apo-E receptor binding. Evidence from the analysis of plasma lipoprotein patterns and tissue culture reactivity suggested that changes in assayed hepatic lipoprotein receptor activity occurred in concert with changes in plasma lipoproteins.The results indicate that the two canine hepatic lipoprotein receptors differ in their metabolic regulation. The apo-B,E receptor responds rapidly to changes in hepatic requirements for cholesterol. The apo-E receptor appears to be more refractory to acute regulation. The rapidity of the changes in the activity of the apo-B,E receptor (within 2-4 h) suggests that the binding activity of this receptor may be regulated by factors independent of protein synthesis.

The present investigation describes the influence of partial, pharmacologic interruption of bile acid enterohepatic circulation on cholesterol and bile acid synthesis in germfree rats. Seven rats received a basal, semisynthetic diet and five rats received the basal diet supplemented with 5% cholestyramine. After 6 weeks, feces were collected in one 3- and one 4-day pool for analysis of bile acids and cholesterol. When the sampling period was concluded, the rats were killed and the liver microsomal fractions were isolated. The activities of HMG CoA reductase and 7alpha-hydroxylase were determined. The main fecal bile acids in the untreated rats were cholic acid and beta-muricholic acid. During cholestyramine treatment cholic acid increased from 4.4 +/- 0.7 to 39.5 +/- 5.6 mg/kg body weight/day and beta-muricholic acid from 4.5 +/- 0.6 to 7.7 +/- 0.9 mg/kg body weight/day. Chenodeoxycholic acid became a major bile acid averaging 10.4 +/- 1.6 mg/kg body weight/day. The total amount of bile acids increased about 6-7 times and the percentage of cholic acid increased from 49.4 +/- 2.0 to 68.6 +/- 1.1%. The 7alpha-hydroxylase activity increased 4-5 times. During cholestyramine treatment the fecal excretion of cholesterol was increased from 12.0 +/- 1.4 to 68.0 +/- 5.0 mg/kg body weight/day. The endogenous formation of cholesterol was increased 6 times and the HMG CoA reductase activity was increased about 20 times. In conclusion, germfree rats, like conventional rats, have the ability to increase the endogenous formation of bile acids and cholesterol during interruption of the enterohepatic circulation of bile acids, which is also reflected in a stimulation of the activities of the rate-determining enzymes.

The serum concentrations of cholic acid (C), chenodeoxycholic acid (CD), and deoxycholic acid (D) were determined in peripheral venous and portal venous blood from 10 otherwise healthy patients undergoing elective cholecystectomy. A highly specific and accurate gas chromatographic-mass spectrometric technique was used. Peripheral venous serum contained 0.49 +/- 0.16 (mean +/- SEM) mumole per liter of C, 1.55 +/- 0.32 mumoles per liter of CD, and 1.44 +/- 0.57 mumoles per liter of D. Arterial serum, obtained from 5 of the subjects, did not show any differences in bile acid concentrations compared to venous serum. In contrast, the portal venous content of each bile acid was several-fold greater, 6.14 +/- 1.20 mumoles per liter of C, 8.40 +/- 1.84 mumoles per liter of CD, and 6.18 +/- 2.27 mumoles per liter of D. The hepatic uptake of C was estimated to be about 90%, whereas that of CD and D was lower, about 70%. This difference in hepatic uptake between the individual bile acids was reflected in the relative composition of the total bile acids, which was 30:39:31 (C:CD:D) in portal venous serum and 13:50:37 in peripheral serum. Compared to common duct bile obtained simultaneously, the portal vein contained a greater proportion of CD. The relevance of the data obtained to our present concept of the enterohepatic circulation of bile acids is discussed, and it is suggested that the higher fasting level of CD compared to C in peripheral serum results from the combination of a lower fractional hepatic extraction and a higher portal venous input to the systemic circulation.

Fasting serum concentrations of cholic acid, chenodeoxycholic acid, and deoxycholic acid were determined in healthy subjects and in patients with familial hypercholesterolemia before and during treatment with cholestyramine. The bile acids were analyzed by a specific isotope-dilution technique by using gas chromatography-mass spectrometry. Cholestyramine treatment did not change the fasting concentration of total bile acids, but the contribution of cholic acid was increased; those of chenodeoxycholic acid and deoxycholic acid were decreased. No decrease of fasting bile-acid concentrations in portal venous serum was seen in 2 cholestyramine-treated gallstone patients. The postprandial total bile-acid concentration was about 40% lower during cholestyramine treatment in healthy subjects, reflecting a reduced postprandial inflow of bile acids to the liver. This degree of interruption of the postprandial enterohepatic circulation may be sufficient to produce a near maximal bile-acid biosynthesis rate and to promote lowering of plasma cholesterol also in the fasting state. It is concluded that the postprandial bile-acid inflow to the liver may be more important as a regulator of bile-acid biosynthesis than is the fasting level of bile acids.